Although the existence of inducible, chromosomally-mediated beta- lactamases has long been recognized, their clinical importance has only recently been appreciated. Loss of control of expression of these enzymes in certain gram-negative bacteria has been responsible for the rapid development of resistance during therapy with many of the newer beta-lactam antibiotics. This resistance affects all of the penicillins, cephalosporins and monobactams. These multiple beta-lactam resistant mutants resultant from the loss of control of chromosomal beta-lactamases are becoming increasingly frequent causes of nosocomial infections especially in areas of the hospital where there is a high density of patients at risk of becoming infected with such organisms. This includes burn units, cancer centers, intensive care units and cystic fibrosis centers. Since the clinical importance of these enzymes has only recently been recognized, much is unknown about these inducible beta- lactamases. Thus, the long range goal of the proposed study is to delineate the control mechanisms responsible for expression of these beta-lactamases in Enterobacter cloacae and Pseudomonas aeruginosa -- two organisms that commonly cause nosocomial infections. Only from such information can approaches to prevent loss of control be designed.
Two specific aims will be directed at accomplishing this goal: (i) Delineation of the genetic organization responsible for the enzymes. Genetic regions will be cloned and transferred to suitable recipients to identify both control and structural genes; and (ii) Examination of the impact of control and structural genes on the physiology of the cell. Cellular proteins and processes will be examined to determine the impact of the genes identified in Specific Aim 1 upon synthesis of both beta- lactamase and cell wall. This should allow the assessment of the more global impact of the genes on cellular physiology. Major methods for accomplishing these aims will include electrophoresis, isoelectric focusing, spectrophotometric assays for beta-lactamase, and use of various recombinant DNA techniques for development of gene probes, cloning, transfer of DNA, allelic replacement, and replacement mutagenesis.
| Sanders, C C; Bradford, P A; Ehrhardt, A F et al. (1997) Penicillin-binding proteins and induction of AmpC beta-lactamase. Antimicrob Agents Chemother 41:2013-5 |
| Pitout, J D; Moland, E S; Sanders, C C et al. (1997) Beta-lactamases and detection of beta-lactam resistance in Enterobacter spp. Antimicrob Agents Chemother 41:35-9 |
| Ehrhardt, A F; Sanders, C C; Romero, J R et al. (1996) Sequencing and analysis of four new Enterobacter ampD Alleles. Antimicrob Agents Chemother 40:1953-6 |
